title: 3 beta-hydroxysteroid oxidoreductase in suspension cultures of Digitalis lanata EHRH.
abstract: A 3 beta-hydroxysteroid oxidoreductase was isolated and characterized in the microsomes of Digitalis lanata cell cultures. The enzyme catalyzes the conversion of 5 alpha-pregnane-3,20-dione to 5 alpha-pregnan-3 beta-ol-20-one and requires NAD(P)H2. The enzyme was found to have a pH optimum of 8.0. The reaction had an optimum incubation temperature of 25 degrees C with linear reduction for the first 4 h, reaching maximum enzyme activity after 7 h. Substrate kinetics for 5 alpha-pregnane-3,20-dione and NADPH2 resulted in apparent Km-values of 18.5-20 microM for 5 alpha-pregnane-3,20-dione and 50-120 microM for the co-substrate NADPH2. In order to localize 3 beta-hydroxysteroid oxidoreductase differential centrifugation as well as linear sucrose density gradient centrifugation were performed. The results obtained lead to the conclusion that 3 beta-hydroxysteroid oxidoreductase is not associated with a single cell compartment, but consists of a major soluble part and a markedly smaller part of endoplasmic reticulum-associated activity.
authors.affiliation: Universität Tübingen, Botanisches Institut, Allgemeine Botanik und Pflanzenphysiologie, Bundesrepublik Deutschland.
authors.forename: H M
authors.initials: HM
authors.lastname: Warneck
authors.forename: H U
authors.initials: HU
authors.lastname: Seitz
mesh_terms: D015096:3-Hydroxysteroid Dehydrogenases / Q000302:isolation & purification / Q000378:metabolism*; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D004070:Digitalis / Q000201:enzymology*; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D010946:Plants, Medicinal; D010947:Plants, Toxic
chemical_list: D015096:3-Hydroxysteroid Dehydrogenases